Can't exec "samtools"

[jxcao98]

Hi

Thanks for your nice tool!

I tried to run VEP by Singularity, but I got confused about where the path of Samtools should be placed.

I have added the path of Samtools to ~/.bashrc. However, I still got a warning, and It could appear multiple times.

Can't exec "samtools": No such file or directory at /home1/GENE_proc/tools/ensemblVEP/Plugin/LoF.pm line 539, <$fh> line 15248.
Use of uninitialized value $faidx in split at /home1/GENE_proc/tools/ensemblVEP/Plugin/LoF.pm line 540, <$fh> line 15248.
Can't exec "samtools": No such file or directory at /home1/GENE_proc/tools/ensemblVEP/Plugin/LoF.pm line 539, <$fh> line 85248.
Use of uninitialized value $faidx in split at /home1/GENE_proc/tools/ensemblVEP/Plugin/LoF.pm line 540, <$fh> line 85248.

The version of loftee:

[v.1.0.4](https://github.com/konradjk/loftee/releases/tag/v1.0.4_GRCh38)

The following is my script:

singularity exec $VepSif \
	vep \
	--cache --offline --force_overwrite \
	--dir $CacheVEP \
	--assembly GRCh38 \
	--fasta $referenceFastaVEP \
	--input_file $FilePath \
	--format vcf \
	-o stdout \
	--vcf \
	--stats_file $runDir/log/$ChrName.summary.html \
	--warning_file $runDir/log/$ChrName.warning.txt \
	--everything \
	--pick \
	--dir_plugins $PluginPath \
	--plugin dbNSFP,$PluginDataPath/dbnsfp/dbNSFP4.3a_grch38.gz,transcript_match=1,SIFT_pred,Polyphen2_HDIV_pred,Polyphen2_HVAR_pred,LRT_pred,MutationTaster_pred,CADD_phred,FATHMM_pred,MetaSVM_pred,REVEL_score,PROVEAN_pred,M-CAP_pred,PrimateAI_pred \
	--plugin LoF,loftee_path:$PluginPath,human_ancestor_fa:$PluginDataPath/loftee/human_ancestor.fa.gz,gerp_bigwig:$PluginDataPath/loftee/gerp_conservation_scores.homo_sapiens.GRCh38.bw,conservation_file:$PluginDataPath/loftee/loftee.sql \
	--plugin CADD,$PluginDataPath/cadd/GRCh38/whole_genome_SNVs.tsv.gz \
	--plugin Conservation,$PluginDataPath/conservation/gerp_conservation_scores.homo_sapiens.GRCh38.bw \
	| bgzip -c > $runDir/$ChrName.AnnotVEP.vcf.gz

It looked like VEP was not interrupted as I still got the final annotated files. So could I ignore these warnings safely?

Any suggestions are appreciated!

Best,
Jixin

[jon4thin]

Jinxin, was there actually any LoFTEE annotations tho? It probably does not, if the package could not run (it needs samtools). The error says that VEP is looking for samtools at /home1/GENE_proc/tools/ensemblVEP/Plugin/LoF.pm

Make sure you can run samtools from your command line stand alone (try running a samtools command on a vcf).